The production of alcoholic beverages by the fermentation of fruit and grains is of ancient origin. In more recent times, the isolation of ethanol in concentrated or in pure form for use either in beverages, in industry, or as fuel, has assumed considerable importance. In general, ethanol may be produced by the fermentation of simple sugars such as glucose and fructose and oligosaccharides such as sucrose. Such substances and mixtures thereof which, without prior chemical modification, are convertible to ethanol will be referred to herein as “fermentable carbohydrates.” More complex carbohydrates such as starches and cellulosic materials also may be converted to ethanol by fermentation, but usually only after they are degraded to lower molecular weight sugars or related materials. The fermentation proceeds in an anaerobic environment, with production of carbon dioxide by-product. The Kirk-Othmer Encyclopedia of Chemical Technology, Third Edition, John Wiley and Sons, New York, N.Y., presents a condensed summary of the state of the art of producing and isolating fermentation-ethanol in Volume 9, at pages 352-361, which is incorporated herein by reference.
Ethanol has widespread application as an industrial chemical, gasoline additive or straight liquid fuel. As a fuel or fuel additive, ethanol dramatically reduces air emissions while improving engine performance. As a renewable fuel, ethanol reduces national dependence on finite and largely foreign fossil fuel sources while decreasing the net accumulation of carbon dioxide in the atmosphere.
In contrast to energy production by combustion of fossil fuels, energy production by combustion of contemporary biomass (predominantly in the form of harvested plant material) or fuels derived from such biomass is regarded as being “CO2-neutral”, since the amount of CO2 released by combustion of a given amount of such biomass corresponds to the amount of CO2 which was originally taken up from the atmosphere during the build-up of that amount of biomass.
Among fuels derived from plant biomass, ethanol has received particular attention as a potential replacement for or supplement to petroleum-derived liquid hydrocarbon products. To minimize the production cost of ethanol produced from biomass (also referred to in the following as “bioethanol”) it is important to use biomass in the form of low-cost by-products from gardening, agriculture, forestry, the timber industry and the like; thus, for example, materials such as straw, maize stems, forestry waste (log slash, bark, small branches, twigs and the like), sawdust and wood-chips are all materials which can be employed to produce bioethanol.
Biomass typically includes materials containing cellulose, hemicellulose, lignin, protein and carbohydrates such as starch and sugar. Common forms of biomass include trees, shrubs and grasses, corn and corn husks as well as municipal solid waste, waste paper and yard waste. Biomass high in starch, sugar or protein such as corn, grains, fruits and vegetables are usually consumed as food. Conversely, biomass high in cellulose, hemicellulose and lignin are not readily digestible and are primarily utilized for wood and paper products, fuel, or are disposed of.
Biomass contains two basic constituents, carbohydrates and lignin. The carbohydrate content of the biomass contains cellulose and hemicellulose. Both cellulose and hemicellulose can be converted to sugars of glucose and xylose. Fermentation converts glucose and xylose to ethanol using enzymes produced by microorganisms, for example, as shown in U.S. Pat. No. 5,789,210. Control of nutrients, pH, temperature, sugar concentration, and microorganism concentration all affect rate of fermentation to form ethanol. When ethanol concentration reaches above about 6 to 12%, ethanol concentration can be lethal to the microorganisms employed for fermentation. To reduce ethanol concentration within broth employed for fermentation and maintain activity of microorganisms, extraction of ethanol from the broth by solvents non-toxic to microorganisms, as disclosed in U.S. Pat. Nos. 5,110,319, 4,865,973 and No. 4,517,298. The operations disclosed require energy for vaporization of ethanol and subsequent condensation to produce liquid ethanol.
The various operations in processes for obtaining ethanol from such recurring sources as cellulose, cane sugar, amylaceous grains and tubers, e.g., the separation of starch granules from non-carbohydrate plant matter and other extraneous substances, the acid and/or enzymatic hydrolysis of starch and/or cellulose to fermentable sugar (saccharification), the fermentation of sugar to a dilute solution of ethanol and the recovery of anhydrous ethanol by distillation, have been modified in numerous ways to achieve improvements in product yield, production rates and so forth. For ethanol to realize its vast potential as a partial or total substitute for petroleum fuels or as a substitute chemical feedstock, it is necessary that the manufacturing process be as efficient in the use of energy and raw materials as possible so as to maximize the energy return for the amount of ethanol produced and enhance the standing of the ethanol as an economically viable replacement for petroleum-based chemicals. To date, however, relatively little concern has been given to the energy requirements for manufacturing ethanol from biomass and consequently, little effort has been made to minimize the thermal expenditure for carrying out any of the discrete operations involved in the manufacture of ethanol from vegetative sources.
There are three basic methods known for accelerating the fermentation rates of a sugar media to ethanol. These methods include 1) increasing the cell density, and/or 2) reducing the concentration of inhibitory compound(s) (with ethanol being most inhibitory due to its osmolality and toxic effects) in the media as suggested by Dale in prior U.S. Pat. Nos. 4,665,027 and 5,141,861. The third method is to control the growth environment; such as with trace elements, vitamins, amino acid, pH, and temperature (Biology of Microorganisms, 7th edition, Brock, Madigan, Martinko and Parler. Prenitice Hall, Ney Jersey 1994). During a normal batch ethanol fermentation with standard S. cerevisae strains, a final cell concentration of between 1.5 and 15 g/l cells is achieved. It is often noted that cell growth completely stops after a certain cell density is reached (Holzberg et al, 1967). The oxygen tension in the fermentation is important in these batch fermentations, as the cells will convert a larger fraction of the sugar substrate towards cell mass production as the amount of oxygen available to the cells increases. Trace oxygen can serve as a nutrient during the anaerobic fermentation of sugars, allowing more cell production that results in a greater fermentation rate of sugar to ethanol. Cysewski and Wilke (1978) show an optimal oxygen tension of about 0.1 mm O2. To maintain a cell density higher than the natural maximum attained in the fermenter, methods for keeping the cells in the fermenter must be utilized. A high cell density can be maintained either by recycling cells (through membrane or centrifugal techniques) or by retaining or immobilizing the cells within the reactor. Immobilization would seem to be advantageous as the capital expense of a cell recovery and recycle system can be eliminated. There has been a good deal of work over the last 10-15 years on immobilizing organisms to maintain a high cell density in the bioreactor. Immobilization can take one of several approaches, 1) entrapment within a gel bead or plate, 2) adsorption onto a solid matrix, or 3) self-agglomeration or flocculation into flakes or pellets.
Increasing temperature generally speeds a fermentation until the temperature becomes high enough to cause cell death. Fermentation rates are generally noted to increase from 20° to 32° C., doubling with a 5° C. increase in temperature.
The traditional process of fermentation is carried out in a conventional batch operation utilizing yeast as the fermenting organism. To increase the efficiency a variation of the batch operation occasionally includes recycling of the yeast cells by systems such as sedimentation, centrifugation, or ultrafiltration. Normally this batch operation is conducted in two stages. The first stage involves propagation of the yeast and is referred to as the growth stage. The second stage involves the anaerobic process of ethanol production which is accompanied by a depletion of the oxygen. Further propagation of the yeast occurs during the anaerobic process of ethanol production.
Typically, a yeast inoculum is prepared in stage one. The requirements for maximum yeast reproduction are adequate amounts of carbon, nitrogen, minerals and oxygen, a pH in the range of 3.5 to 4.5, and a temperature in the range of 29°-35° C. Aerobic growth conditions define a system for more efficient production of yeast but under which no ethanol is produced. Stage two is the fermentation stage where the alcohol is actually produced by the yeast from the fermentable sugars. The yeast inoculum produced in stage one is used to seed a large fermenter containing glucose at appropriate pH, temperature and sugar concentration. Glucose was formed from the conversion of dextrins via saccharification enzymes. The dextrins were derived from molasses, corn starch materials, etc. via liquification enzymes. The inoculation rate can be 5 to 10 million cells per ml and during the fermentation the viable cell count can increase to 150-200 million cells per ml. Heat produced is controlled through the use of cooling coils. At these yeast levels, a final ethanol concentration of about 9 to 11% (v/v) can be obtained in 30 to 70 hours with batch fermentation. Increasing the yeast content, as is the case with cell recycle, can considerably reduce the time required for completion of the fermentation. For example, with a cell density of 800 to 1000 million cells per ml, it is possible to reduce the fermentation time to 4 to 10 hours.
Processes for the continuous fermentation of sugars to provide alcohol are also well known (U.S. Pat. Nos. 2,155,134; 2,371,208; 2,967,107; 3,015,612; 3,078,166; 3,093,548; 3,177,005; 3,201,328; 3,207,605; 3,207,606; 3,219,319; 3,234,026; 3,413,124; 3,528,887; 3,575,813; 3,591,454; 3,705,841; 3,737,323; and 3,940,492 “Process Design and Economic Studies of Alternative Fermentation Methods for the Production of Ethanol”, Cysewski, et al. Biotechnology and Bioengineering, Vol. xx, pp. 1421-1444 (1978)). In a typical continuous fermentation process, a stream of sterile sugar liquor and a quantity of yeast cells are introduced into the first of a battery of fermentation vessels wherein initial fermentation takes place, generally under conditions favoring rapid cell growth. The partial fermentate admixed with yeast cells is continuously withdrawn from the first fermentation vessel wherein fermentation is carried out under conditions favoring the rapid conversion of sugar to ethanol. The yeast in the last fermentation vessel can be recovered by suitable means, e.g., centrifugation or settlement, and recycled. In such a system, the ability of the fermentation organism to produce ethanol is affected by the ethanol and sugar concentrations. As a rule, a yeast which gives high conversion rates of sugar to ethanol in a low-ethanol, high-sugar fermentation medium will only sluggishly produce ethanol under the opposite conditions, i.e., at high-ethanol level, low-sugar concentrations.
In general, however, the price of bioethanol has not been competitive with that of traditional fossil fuels and it is therefore highly needed to reduce production costs as far as possible by optimizing or improving upon bioethanol production technologies.